Application Note: Nonlinear Microscopy

By IMRA America, Inc.

Ultrafast lasers have revolutionized the field of microscopy by enabling new types of nonlinear microscopy techniques. Femtosecond pulse lasers provide the combination of high peak power (1-100 kW) and low average power (10-1000 mW) which is ideal for utilizing nonlinear optical processes in scanning laser microscopy (LSM). The majority of LSM work is done using normal, single-photon fluorescence using blue and green laser sources. However, nonlinear microscopy, offers certain advantages over single-photon methods: (1) Natural sectioning without the need for a confocal de-scanned aperture; (2) greater penetration depth in scattering media due to use of longer wavelengths, and; (3) Reduced photo-toxicity due to smaller excitation volume, resulting in longer viewing time of living cells and organisms. It is these advantages which explain why multiphoton LSM is enjoying growing popularity among researchers in biology, medicine, and neuroscience. Here, we describe the two main varieties of nonlinear microscopy: two-photon microscopy, and third-harmonic generation (THG) microscopy. And we show images using these techniques with various types of modelocked fiber lasers.